The reaction to injury hypothesis postulates that the initiating factor in the induction of pathologic migration and proliferation of smooth muscle (SMC) from the media to the intima is subtle or frank endothelial injury or dysfunction. Although the initiation and development of the lesion is probably multifactoral, involving the participation of many cell types, the initial stimulus to the SMC may be derived from adherence and degranulation of platelets, extravasation of macrophages into the subendothelial space, or from endothelial cells. Endothelial cells in vitro secrete at least two mitogens, capable of supporting SMC migration and proliferation. One mitogen is similar or identical to platelet-derived growth factor (PDGF). The largest portion of mitogenic activity is distinct from PDGF in receptor competition assays, behavior during ion exchange chromatography, and isoelectric points. Information on the control of mitogen secretion by endothelium is beginning to emerge. The PDGF mitogen is expressed only at extremely low levels under normal in vivo conditions but is constitutively secreted in vitro. This project will investigate the conditions that induce endothelium to express PDGF during in vivo to in vitro transfer and conditions that perturb secretion in vitro using various model systems. Three probes will be used: competition for PDGF receptors, mitogenesis assays, and hybridization with c-sis cDNA. The second area of focus is the regulation of nonPDGF mitogen secretion. This mitogen is specifically released from endothelium by heparin, a putative inhibitor of SMC migration and proliferation. Proposed experiments will investigate how heparin and growth factor interact with the endothelium and with each other and how the endothelium functions as a secretory cell, responding to appropriate stimuli by secreting large amounts of intracellulr nonPDGF mitogen. The third aim of this proposal is purification of these mitogens, with emphasis on the nonPDGF mitogens, by using ion exchange, affinity and gel permeation chromatography and HPLC, and generation of specific antibodies. It is critical to understand the conditions and the circumstances that regulate endothelial cell mitogen secretion and the conditions that promote or induce expression of these activities to understand the role that the endothelium plays in induction or prevention of the disease process.